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CADDIS Volume 3: Examples & Applications

Analytical Examples

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Analytical techniques used:

Type of evidence supported:

Stressor-Response Relationships from Laboratory Studies


In this example, we ask whether organisms in Long Creek, Maine (U.S. EPA 2007) are exposed to a candidate cause (zinc) at quantities or frequencies sufficient to induce observed biological effects. We use results from laboratory studies to evaluate whether zinc in the water column under base flow conditions reached concentrations that could explain the observed decrease in Ephemeroptera, Plecoptera and Trichoptera (EPT) richness. The comparison of laboratory and field data can be performed in two ways.

  1. Most commonly, effective concentrations from laboratory data are compared to ambient concentrations at the affected site. If zinc concentrations associated with similar types of effects in the laboratory are similar to or lower than concentrations that have been shown to occur at the affected site, this would provide evidence that zinc concentrations are high enough to cause the effects. Conversely, if zinc concentrations associated with similar types of effects in the laboratory are much higher than those at the affected site, then the case for zinc would be weakened. Either some other stressor is the cause of the observed decline, or zinc is acting jointly with another cause to produce the effect.

  2. We can also compare the magnitude of effects observed at the site with the magnitude of effects observed in the laboratory at concentrations equal to ambient concentrations. If the magnitude of effects at the site are much greater than would be predicted from the laboratory concentration-response relationship, then we would conclude that either zinc concentrations are not high enough to have caused the effects, or the laboratory organisms or endpoints are not as sensitive as the organisms or responses at the affected site. If magnitude of effects observed at the site is approximately equal to those predicted from the laboratory concentration-response relationship, then this would support the argument that zinc is the cause of the effects. Finally, if the magnitude of effects observed at the site is much less than predicted from laboratory studies, we would conclude that some physical factor (e.g., dissolved organic matter) or some biological process (e.g., replacement of sensitive insect species by tolerant species) may be reducing the effect in the field.


This example uses summaries of laboratory toxicity test results and compares these summaries with data from the site.

Laboratory toxicity data

Two approaches were used to summarize laboratory results. First, U.S. EPA's chronic criterion value for zinc was used to represent sublethal effects and effects of extended exposures. The chronic criterion value for zinc at 100 mg/L hardness (as CaCO3) is 0.12 mg/L. A chronic value for an EPT insect would be preferable, but none were available.

Second, species sensitivity distributions (SSDs) were developed using data from the ECOTOX database. The project team selected freshwater aquatic organism tests with site-appropriate water hardness, pH and temperature. Data were further subdivided to generate SSDs addressing potential effects at baseflow/lowflow exposure (3-30 days) and at stormflow/pulsed exposures (<30 hours).

It was necessary to generate SSDs with data for total metals because greater than 90% of freshwater metals data in ECOTOX are reported as total metals. Free ion or dissolved metal concentrations would be more appropriate indicators of actual toxic exposure and be more relevant to the dissolved metal concentrations reported for Long Creek. However, this is a relatively minor problem, because nearly all metals in laboratory tests are dissolved.

SSDs were generated using LC50 data. Since an LC50 is a concentration that kills half of the organisms in a test population, one would expect to observe a reduction in the abundance of some species when water concentrations equal the LC50 for that species. Data used in generating SSDs do not represent specific species present at the study area. Toxicity data are generally not available for site-specific taxa due to the diversity of species occurring in the wild and the need to perform toxicity tests with well characterized organisms.

Site data

Biological and water chemistry data from two sites along Long Creek are used in this example. EPT richness was calculated from macroinvertebrate rockbag samples deployed throughout the study area beginning August 5-6 1999, following standard Maine Department of Environmental Protection (MEDEP) protocol (Davies and Tsomides 2002).

Baseflow water samples were collected by MEDEP on three days in August 2000. Methods and analyses are described in MEDEP (2002).

Analysis and results

The laboratory results were compared to site data by graphically comparing the proportion of decrease in EPT richness, relative to the reference site, and impaired site zinc concentrations. In addition, the SSD was used to identify 0.087 as a benchmark concentration of 10% at which 10% of species would be expected to experience lethal effects.

line graph
Figure 1. Comparison of site observations from Long Creek with the EPA criterion continuous concentration for Zn (EPA CCC) and a species sensitivity distribution. Points A and B mark corresponding biological observations and Zn concentrations from Sites LCMn2.274 and LCN .415, respectively.


The analysis hinges on three assumptions.

  1. The organisms and endpoints measured in the laboratory are relevant to EPT richness.
  2. The laboratory exposures are relevant to the exposures encountered by organisms in the field.
  3. Measured baseflow concentrations of zinc in August 2000 were similar to unmeasured concentrations in August 1999.

How do I score this evidence?

Measured concentrations are all below the EPA criterion continuous concentration. The measured concentrations at the site fall below the 10% benchmark derived from the SSD. Points corresponding to the observed impairment occur at concentrations below the lower confidence limits on the SSD curve. This weakens the case that zinc caused the observed decreases in EPT, giving a score of - (one minus).


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